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1.
Sci Rep ; 14(1): 10926, 2024 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-38740833

RESUMO

In contrast to acute diarrhoea, the aetiology of persistent digestive disorders (≥ 14 days) is poorly understood in low-resource settings and conventional diagnostic approaches lack accuracy. In this multi-country study, we compared multiplex real-time PCR for enteric bacterial, parasitic and viral pathogens in stool samples from symptomatic patients and matched asymptomatic controls in Côte d'Ivoire, Mali and Nepal. Among 1826 stool samples, the prevalence of most pathogens was highest in Mali, being up to threefold higher than in Côte d'Ivoire and up to tenfold higher than in Nepal. In all settings, the most prevalent bacteria were EAEC (13.0-39.9%) and Campylobacter spp. (3.9-35.3%). Giardia intestinalis was the predominant intestinal protozoon (2.9-20.5%), and adenovirus 40/41 was the most frequently observed viral pathogen (6.3-25.1%). Significantly different prevalences between symptomatic and asymptomatic individuals were observed for Campylobacter, EIEC and ETEC in the two African sites, and for norovirus in Nepal. Multiple species pathogen infection was common in Côte d'Ivoire and Mali, but rarely found in Nepal. We observed that molecular testing detected multiple enteric pathogens and showed low discriminatory accuracy to distinguish between symptomatic and asymptomatic individuals. Yet, multiplex PCR allowed for direct comparison between different countries and revealed considerable setting-specificity.


Assuntos
Dor Abdominal , Diarreia , Fezes , Reação em Cadeia da Polimerase Multiplex , Humanos , Côte d'Ivoire/epidemiologia , Diarreia/microbiologia , Diarreia/parasitologia , Diarreia/virologia , Diarreia/epidemiologia , Diarreia/diagnóstico , Reação em Cadeia da Polimerase Multiplex/métodos , Nepal/epidemiologia , Mali/epidemiologia , Masculino , Feminino , Adulto , Fezes/microbiologia , Fezes/parasitologia , Fezes/virologia , Adolescente , Criança , Pessoa de Meia-Idade , Pré-Escolar , Adulto Jovem , Lactente , Prevalência , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/classificação , Idoso , Giardia lamblia/isolamento & purificação , Giardia lamblia/genética
2.
Parasitol Res ; 123(4): 179, 2024 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-38584235

RESUMO

Giardia duodenalis, the protozoan responsible for giardiasis, is a significant contributor to millions of diarrheal diseases worldwide. Despite the availability of treatments for this parasitic infection, therapeutic failures are alarmingly frequent. Thus, there is a clear need to identify new therapeutic targets. Giardia telomeres were previously identified, but our understanding of these structures and the critical role played by Giardia telomerase in maintaining genomic stability and its influence on cellular processes remains limited. In this regard, it is known that all Giardia chromosomes are capped by small telomeres, organized and protected by specific proteins that regulate their functions. To counteract natural telomere shortening and maintain high proliferation, Giardia exhibits constant telomerase activity and employs additional mechanisms, such as the formation of G-quadruplex structures and the involvement of transposable elements linked to telomeric repeats. Thus, this study aims to address the existing knowledge gap by compiling the available information (until 2023) about Giardia telomeres and telomerase, focusing on highlighting the distinctive features within this parasite. Furthermore, the potential feasibility of targeting Giardia telomeres and/or telomerase as an innovative therapeutic strategy is discussed.


Assuntos
Giardia lamblia , Giardíase , Telomerase , Humanos , Telomerase/genética , Telomerase/metabolismo , Giardíase/parasitologia , Giardia/genética , Telômero/genética , Giardia lamblia/genética , Giardia lamblia/metabolismo
3.
Parasitol Res ; 123(4): 176, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38573530

RESUMO

Giardiasis is a common intestinal infection caused by Giardia duodenalis, which is a major economic and health burden for humans and livestock. Currently, a convenient and effective detection method is urgently needed. CRISPR/Cas12a-based diagnostic methods have been widely used for nucleic acid-based detection of pathogens due to their high efficiency and sensitivity. In this study, a technique combining CRISPR/Cas12a and RPA was established that allows the detection of G. duodenalis in faecal samples by the naked eye with high sensitivity (10-1 copies/µL) and specificity (no cross-reactivity with nine common pathogens). In clinical evaluations, the RPA-CRISPR/Cas12a-based detection assay detected Giardia positivity in 2% (1/50) of human faecal samples and 47% (33/70) of cattle faecal samples, respectively, which was consistent with the results of nested PCR. Our study demonstrated that the RPA-CRISPR/Cas12a technique for G. duodenalis is stable, efficient, sensitive, specific and has low equipment requirements. This technique offers new opportunities for on-site detection in remote and poor areas.


Assuntos
Giardia lamblia , Giardíase , Humanos , Animais , Bovinos , Giardia lamblia/genética , Sistemas CRISPR-Cas , Giardíase/diagnóstico , Giardíase/veterinária , Giardia/genética , Bioensaio
4.
Int J Biol Macromol ; 267(Pt 2): 131509, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38608978

RESUMO

Giardia intestinalis is one of the most widespread intestinal parasites and is considered a major cause of epidemic or sporadic diarrhea worldwide. In this study, we aimed to develop a rapid aptameric diagnostic technique for G. intestinalis infection. First, the SELEX (Systematic Evolution of Ligands by Exponential Enrichment) process generated DNA aptamers specific to a recombinant protein of the parasite's trophozoite. Ten selection rounds were performed; each round, the DNA library was incubated with the target protein conjugated to Sepharose beads. Then, the unbound sequences were removed by washing and the specific sequences were eluted and amplified by Polymerase Chain Reaction (PCR). Two aptamers were selected, and the dissociation constants (Kd), were determined as 2.45 and 16.95 nM, showed their high affinity for the G. intestinalis trophozoite protein. Subsequently, the aptamer sequence T1, which exhibited better affinity, was employed to develop a label-free electrochemical biosensor. A thiolated aptamer was covalently immobilized onto a gold screen-printed electrode (SPGE), and the binding of the targeted protein was monitored using square wave voltammetry (SWV). The developed aptasensor enabled accurate detection of the G. intestinalis recombinant protein within the range of 0.1 pg/mL to 100 ng/mL, with an excellent sensitivity (LOD of 0.35 pg/mL). Moreover, selectivity studies showed a negligible cross-reactivity toward other proteins such as bovine serum albumin, globulin, and G. intestinalis cyst protein.


Assuntos
Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , Técnicas Eletroquímicas , Giardia lamblia , Proteínas de Protozoários , Técnica de Seleção de Aptâmeros , Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Técnica de Seleção de Aptâmeros/métodos , Técnicas Eletroquímicas/métodos , Proteínas de Protozoários/química , DNA de Cadeia Simples/química , Giardíase/diagnóstico , Giardíase/parasitologia
5.
Front Public Health ; 12: 1377123, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38645455

RESUMO

Introduction: Giardiosis remains one of the most prevalent enteric parasitic infections globally. Earlier molecular-based studies conducted in Egypt have primarily focused on paediatric clinical populations and most were based on single genotyping markers. As a result, there is limited information on the frequency and genetic diversity of G. duodenalis infections in individuals of all age groups. Methods: Individual stool samples (n = 460) from outpatients seeking medical care were collected during January-December 2021 in Kafr El-Sheikh governorate, northern Egypt. Initial screening for the presence of G. duodenalis was conducted by coprological examination. Microscopy-positive samples were further confirmed by real-time PCR. A multilocus sequence typing approach targeted amplification of the glutamate dehydrogenase (gdh), beta-giardin (bg), and triose phosphate isomerase (tpi) genes was used for genotyping purposes. A standardised epidemiological questionnaire was used to gather basic sociodemographic and clinical features of the recruited patients. Results: Giardia duodenalis cysts were observed in 5.4% (25/460, 95% CI: 3.6-7.9) of the stool samples examined by conventional microscopy. The infection was more frequent in children under the age of 10 years and in individuals presenting with diarrhoea but without reaching statistical significance. Stool samples collected during the winter period were more likely to harbour G. duodenalis. All 25 microscopy-positive samples were confirmed by real-time PCR, but genotyping data was only available for 56.0% (14/25) of the isolates. Sequence analyses revealed the presence of assemblages A (78.6%, 11/14) and B (21.4%, 3/14). All assemblage A isolates were identified as sub-assemblage AII, whereas the three assemblage B sequences belonged to the sub-assemblage BIII. Patients with giardiosis presenting with diarrhoea were more frequently infected by the assemblage A of the parasite. Conclusion: This is one of the largest epidemiological studies evaluating G. duodenalis infection in individuals of all age groups in Egypt. Our molecular data suggest that G. duodenalis infections in the surveyed population are primarily of anthropic origin. However, because assemblages A and B are zoonotic, some of the infections identified can have an animal origin. Additional investigations targeting animal (domestic and free-living) and environmental (water) samples are warranted to better understand the epidemiology of giardiosis in Egypt.


Assuntos
Fezes , Giardia lamblia , Giardíase , Pacientes Ambulatoriais , Humanos , Egito/epidemiologia , Giardíase/epidemiologia , Feminino , Masculino , Giardia lamblia/genética , Giardia lamblia/isolamento & purificação , Criança , Fezes/parasitologia , Adulto , Pré-Escolar , Adolescente , Pacientes Ambulatoriais/estatística & dados numéricos , Adulto Jovem , Microscopia , Pessoa de Meia-Idade , Tipagem de Sequências Multilocus , Lactente , Genótipo , Reação em Cadeia da Polimerase em Tempo Real
6.
Structure ; 32(4): 377-379, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38579678

RESUMO

Eiler et al. used cryo-electron microscopy to determine a 2.49 Å resolution structure of Giardia lamblia 80S ribosome bound to tRNA, mRNA, and the anti-protozoal drug emetine. The structure reveals some critical aspects of translation in G. lamblia, including the lack of ribosomal protein RACK1, and how emetine blocks translation by interacting with both the ribosome and mRNA.


Assuntos
Giardia lamblia , Giardia lamblia/química , Giardia lamblia/genética , Giardia lamblia/metabolismo , Microscopia Crioeletrônica , Emetina/metabolismo , Ribossomos/metabolismo , RNA Mensageiro/metabolismo
7.
PLoS Pathog ; 20(4): e1012140, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38598600

RESUMO

The Giardia lamblia virus (GLV) is a non-enveloped icosahedral dsRNA and endosymbiont virus that infects the zoonotic protozoan parasite Giardia duodenalis (syn. G. lamblia, G. intestinalis), which is a pathogen of mammals, including humans. Elucidating the transmission mechanism of GLV is crucial for gaining an in-depth understanding of the virulence of the virus in G. duodenalis. GLV belongs to the family Totiviridae, which infects yeast and protozoa intracellularly; however, it also transmits extracellularly, similar to the phylogenetically, distantly related toti-like viruses that infect multicellular hosts. The GLV capsid structure is extensively involved in the longstanding discussion concerning extracellular transmission in Totiviridae and toti-like viruses. Hence, this study constructed the first high-resolution comparative atomic models of two GLV strains, namely GLV-HP and GLV-CAT, which showed different intracellular localization and virulence phenotypes, using cryogenic electron microscopy single-particle analysis. The atomic models of the GLV capsids presented swapped C-terminal extensions, extra surface loops, and a lack of cap-snatching pockets, similar to those of toti-like viruses. However, their open pores and absence of the extra crown protein resemble those of other yeast and protozoan Totiviridae viruses, demonstrating the essential structures for extracellular cell-to-cell transmission. The structural comparison between GLV-HP and GLV-CAT indicates the first evidence of critical structural motifs for the transmission and virulence of GLV in G. duodenalis.


Assuntos
Giardia lamblia , Giardiavirus , Giardia lamblia/ultraestrutura , Giardia lamblia/patogenicidade , Giardiavirus/genética , Microscopia Crioeletrônica , Animais , Capsídeo/ultraestrutura , Capsídeo/metabolismo , Humanos , Filogenia
8.
Talanta ; 274: 126000, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38608630

RESUMO

Luminescent ß-diketonate-europium(III) complexes have been found a wide range of applications in time-gated luminescence (TGL) bioassays, but their poor water solubility is a main problem that limits their effective uses. In this work we propose a simple and general strategy to enhance the water solubility of luminescent ß-diketonate-europium(III) complexes that permits facile synthesis and purification. By introducing the fluorinated carboxylic acid group into the structures of ß-diketone ligands, two highly water-soluble and luminescent Eu3+ complexes, PBBHD-Eu3+ and CPBBHD-Eu3+, were designed and synthesized. An excellent solubility exceeding 20 mg/mL for PBBHD-Eu3+ was found in a pure aqueous buffer, while it also displayed strong and long-lived luminescence (quantum yield φ = 26%, lifetime τ = 0.49 ms). After the carboxyl groups of PBBHD-Eu3+ were activated, the PBBHD-Eu3+-labeled streptavidin-bovine serum albumin (SA-BSA) conjugate was prepared, and successfully used for the immunoassay of human α-fetoprotein (AFP) and the imaging of an environmental pathogen Giardia lamblia under TGL mode, which demonstrated the practicability of PBBHD-Eu3+ for highly sensitive TGL bioassays. The carboxyl groups of PBBHD can also be easily derivatized with other reactive chemical groups, which enables PBBHD-Eu3+ to meet diverse requirements of biolabeling technique, to provide new opportunities for developing functional europium(III) complex biolabels serving for TGL bioassays.


Assuntos
Európio , Solubilidade , Água , Európio/química , Água/química , Humanos , Medições Luminescentes/métodos , Soroalbumina Bovina/química , Complexos de Coordenação/química , Complexos de Coordenação/síntese química , Giardia lamblia/efeitos dos fármacos , Luminescência , Animais , Bioensaio/métodos , Substâncias Luminescentes/química , Substâncias Luminescentes/síntese química , Estreptavidina/química , Fatores de Tempo , Bovinos , Cetoácidos/química
9.
Eur J Protistol ; 93: 126066, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38442435

RESUMO

The zoonotic potential of the protist parasites Cryptosporidium spp. and Giardia duodenalis in amphibians and reptiles raises public health concerns due to their growing popularity as pets. This review examines the prevalence and diversity of these parasites in wild and captive amphibians and reptiles to better understand the zoonotic risk. Research on Giardia in both groups is limited, and zoonotic forms of Cryptosporidium or Giardia have not been reported in amphibians. Host-adapted Cryptosporidium species dominate in reptiles, albeit some reptiles have been found to carry zoonotic (C. hominis and C. parvum) and rodent-associated (C. tyzzeri, C. muris and C. andersoni) species, primarily through mechanical carriage. Similarly, the limited reports of Giardia duodenalis (assemblages A, B and E) in reptiles may also be due to mechanical carriage. Thus, the available evidence indicates minimal zoonotic risk associated with these organisms in wild and captive frogs and reptiles. The exact transmission routes for these infections within reptile populations remain poorly understood, particularly regarding the importance of mechanical carriage. Although the risk appears minimal, continued research and surveillance efforts are necessary to gain a more comprehensive understanding of the transmission dynamics and ultimately improve our ability to safeguard human and animal health.


Assuntos
Criptosporidiose , Cryptosporidium , Giardia lamblia , Giardíase , Animais , Humanos , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Zoonoses/epidemiologia , Zoonoses/parasitologia , Anuros , Répteis , Prevalência , Fezes/parasitologia
10.
Parasitol Res ; 123(3): 160, 2024 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-38480554

RESUMO

Dog faecal samples examined from January 2019 to December 2019 were retrospectively analysed for frequency of endoparasites. The examinations were performed with several different methods: 29,219 samples were examined by flotation method and sodium acetate-acetic acid-formalin concentration (SAFC) technique, 1,330 samples by Baermann-Wetzel migration technique, 12,221 samples using a Giardia coproantigen enzyme-linked-immunosorbent assay (ELISA), 1,180 samples using a Cryptosporidium coproantigen ELISA, 1,671 samples by polymerase chain reaction (PCR) testing for Giardia duodenalis and 447 samples by PCR testing for Cryptosporidium spp.. A total of 7.1% of the samples were positive for parasites in the microscopical examination using the flotation method and SAFC technique. The parasites found included Cystoisospora spp. (2.8%), Giardia duodenalis (2.3%), Ancylostomatidae (1.8%), Toxocara canis (1.6%), Trichuris vulpis (0.7%), Toxascaris leonina (0.5%), Capillaria spp. (0.2%), Angiostrongylus vasorum (0.2%), Crenosoma vulpis (0.1%), Taeniidae (0.1%), Sarcocystis spp. (0.03%), Dipylidium caninum (0.01%), Diphyllobothrium latum (< 0.01%), Spirurida (< 0.01%) and Opisthorchiidae (< 0.01%). Using the Baermann-Wetzel migration technique, Angiostrongylus vasorum was found in 0.75% and Crenosoma vulpis in 0.3% of the samples. ELISAs for Giardia duodenalis and Cryptosporidium spp. revealed 13.9% and 1.0% positive faecal samples, and Giardia duodenalis and Cryptosporidium spp. PCRs 19.4% and 2.0%, respectively. Dogs in the first year of life were more frequently infected with parasites than older animals. In the microscopic examination using flotation method and SAFC technique, the significantly highest detection rates were found in dogs up to six months of age (p < 0.001).


Assuntos
Angiostrongylus , Criptosporidiose , Cryptosporidium , Doenças do Cão , Giardia lamblia , Enteropatias Parasitárias , Parasitos , Cães , Animais , Enteropatias Parasitárias/parasitologia , Estudos Retrospectivos , Doenças do Cão/diagnóstico , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Prevalência , Alemanha/epidemiologia , Fezes/parasitologia
11.
Nanoscale ; 16(14): 7145-7153, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38502112

RESUMO

The unicellular parasite Giardia duodenalis is the causative agent of giardiasis, a gastrointestinal disease with global spread. In its trophozoite form, G. duodenalis can adhere to the human intestinal epithelium and a variety of other, artificial surfaces. Its attachment is facilitated by a unique microtubule-based attachment organelle, the so-called ventral disc. The mechanical function of the ventral disc, however, is still debated. Earlier studies postulated that a dynamic negative pressure under the ventral disc, generated by persistently beating flagella, mediates the attachment. Later studies suggested a suction model based on structural changes of the ventral discs, substrate clutching or grasping, or unspecific contact forces. In this study, we aim to contribute to the understanding of G. duodenalis attachment by investigating detachment characteristics and determining adhesion forces of single trophozoites on a smooth glass surface (RMS = 1.1 ± 0.2 nm) by fluidic force microscopy (FluidFM)-based single-cell force spectroscopy (SCFS). Briefly, viable adherent trophozoites were approached with a FluidFM micropipette, immobilized to the micropipette aperture by negative pressure, and detached from the surface by micropipette retraction while retract force curves were recorded. These force curves displayed novel and so far undescribed characteristics for a microorganism, namely, gradual force increase on the pulled trophozoite, with localization of adhesion force shortly before cell detachment length. Respective adhesion forces reached 7.7 ± 4.2 nN at 1 µm s-1 pulling speed. Importantly, this unique force pattern was different from that of other eukaryotic cells such as Candida albicans or oral keratinocytes, considered for comparison in this study. The latter both displayed a force pattern with force peaks of different values or force plateaus (for keratinocytes) indicative of breakage of molecular bonds of cell-anchored classes of adhesion molecules or membrane components. Furthermore, the attachment mode of G. duodenalis trophozoites was mechanically resilient to tensile forces, when the pulling speeds were raised up to 10 µm s-1 and adhesion forces increased to 28.7 ± 10.5 nN. Taken together, comparative SCSF revealed novel and unique retract force curve characteristics for attached G. duodenalis, suggesting a ligand-independent suction mechanism, that differ from those of other well described eukaryotes.


Assuntos
Giardia lamblia , Giardíase , Animais , Humanos , Giardia lamblia/metabolismo , Trofozoítos/metabolismo , Giardíase/metabolismo , Organelas , Análise Espectral
12.
Acta Trop ; 254: 107188, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38531428

RESUMO

Cryptosporidium spp. and G. duodenalis often infect humans, cats, and other mammals, causing diarrhea and being responsible for numerous outbreaks of waterborne and foodborne infections worldwide. The rapid increase in the number of pet cats poses a substantial public health risk. However, there were few reports about the infection of Cryptosporidium spp. and G. duodenalis infections in pet cats in Henan Province, central China. Thus, to understand the prevalence and genetic distribution of Cryptosporidium spp. and G. duodenalis in pet cats, and to evaluate the zoonotic potential, possible transmission routes and public health implications of isolates, fecal samples (n = 898) were randomly collected from pet cats in 11 cities in Henan Province, central China. Nested PCR based on the SSU rRNA gene and bg gene was used to the prevalence of Cryptosporidium spp. and G. duodenalis, respectively. The prevalence was 0.8 % (7/898) and 2.0 % (18/898) for Cryptosporidium spp. and G. duodenalis respectively. Additionally, the Cryptosporidium spp. positive isolates were identified as C. parvum subtype IIdA19G1 by gp60 gene. In the present study, the IIdA19G1 subtype was discovered in pet cats for the first time in China, enriching the information on the host type and geographical distribution of Cryptosporidium spp. in China. For G. duodenalis, a total of 18 G. duodenalis positive samples were identified, belonging to four assemblages: a zoonotic assemblage A1 (4/898), three host-specific assemblages C (8/898), D (5/898), and F (1/898). Interestingly, we found that pet cats infected with Cryptosporidium spp. and G. duodenalis are more likely to experience emaciation symptoms compared to the negative group. More importantly, the prevalence of Cryptosporidium spp. and G. duodenalis detected in the present study were low, but the subtype IIdA19G1 of Cryptosporidium spp. and the assemblages A1, C, D, and F of G. duodenalis have the potential for zoonotic transmission. Thus, we should focus on preventing and controlling the risk of cross-species transmission that may occur in pet cats in Henan Province.


Assuntos
Doenças do Gato , Criptosporidiose , Cryptosporidium , Fezes , Giardia lamblia , Giardíase , Animais de Estimação , Animais , Gatos , China/epidemiologia , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Criptosporidiose/transmissão , Doenças do Gato/parasitologia , Doenças do Gato/epidemiologia , Cryptosporidium/genética , Cryptosporidium/isolamento & purificação , Cryptosporidium/classificação , Fezes/parasitologia , Giardia lamblia/genética , Giardia lamblia/isolamento & purificação , Giardia lamblia/classificação , Animais de Estimação/parasitologia , Prevalência , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , Giardíase/transmissão , DNA de Protozoário/genética , Filogenia , Reação em Cadeia da Polimerase , Genótipo , Zoonoses/parasitologia , Zoonoses/epidemiologia , Zoonoses/transmissão
13.
Acta Parasitol ; 69(1): 1073-1077, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38499920

RESUMO

PURPOSE: Investigating the genetic variation in thioredoxin reductase (TrxR) and nitroreductase (NR) genes in both treatment-resistant and -sensitive Giardia duodenalis isolates can provide valuable information in identifying potential markers of resistance to metronidazole. The rapid increase in metronidazole treatment failures suggests the presence of genetic resistance mechanisms. By analyzing these genes, researchers can gain insights into the efficacy of metronidazole against G. duodenalis and potentially develop alternative treatment strategies. In this regard, four G. duodenalis isolates (two clinically sensitive and two clinically resistant to metronidazole) were collected from various hospitals of Shiraz, southwestern Iran. METHODS: Parasitological methods including sucrose flotation and microscopy were employed for the primary confirmation of G. duodenalis cysts in stool samples. Microscopy-positive samples were approved by SSU-PCR amplification of the parasite DNA. All four positive G. duodenalis specimens at SSU-PCR were afterward analyzed utilizing designed primers based on important metronidazole metabolism genes including TrxR, NR1, and NR2. RESULTS: Unlike TrxR gene, the results of NR1 and NR2 genes showed that there are non-synonymous variations between sequences of treatment-sensitive and -resistant samples compared to reference sequences. Furthermore, the outcomes of molecular docking revealed that there is an interaction between the protein sequence and spatial shape of treatment-resistant samples and metronidazole in the position of serine amino acid based on the NR1 gene. CONCLUSION: This issue can be one of the possible factors involved in the resistance of Giardia parasites to metronidazole. To reach more accurate results, a large sample size along with simulation and advanced molecular dynamics investigations are needed.


Assuntos
Antiprotozoários , Resistência a Medicamentos , Variação Genética , Giardia lamblia , Giardíase , Metronidazol , Nitrorredutases , Reação em Cadeia da Polimerase , Metronidazol/farmacologia , Giardia lamblia/genética , Giardia lamblia/efeitos dos fármacos , Giardíase/parasitologia , Giardíase/tratamento farmacológico , Humanos , Resistência a Medicamentos/genética , Antiprotozoários/farmacologia , Nitrorredutases/genética , Nitrorredutases/metabolismo , Irã (Geográfico) , Fezes/parasitologia , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Tiorredoxina Dissulfeto Redutase/genética , Tiorredoxina Dissulfeto Redutase/metabolismo , Simulação de Acoplamento Molecular , DNA de Protozoário/genética
14.
Mol Biol Rep ; 51(1): 403, 2024 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-38457002

RESUMO

BACKGROUND: Giardia duodenalis is an important intestinal parasitic protozoan that infects several vertebrates, including humans. Cattle are considered the major source of giardiasis outbreak in humans. This study aimed to investigate the prevalence and multilocus genotype (MLG) of G. duodenalis in Shanxi, and lay the foundation for the prevention and control of Giardiosis. METHODS AND RESULTS: DNA extraction, nested polymerase chain reaction, sequence analysis, MLG analysis, and statistical analysis were performed using 858 bovine fecal samples from Shanxi based on three gene loci: ß-giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi). The overall prevalence of G. duodenalis was 28.3%, while its prevalence in Yingxian and Lingqiu was 28.1% and 28.5%, respectively. The overall prevalence of G. duodenalis in dairy cattle and beef cattle was 28.0% and 28.5%, respectively. G. duodenalis infection was detected in all age groups evaluated in this study. The overall prevalence of G. duodenalis in diarrhea and nondiarrhea samples was 32.4% and 27.5%, respectively, whereas that in intensively farmed and free-range cattle was 35.0% and 19.9%, respectively. We obtained 83, 53, and 59 sequences of bg, gdh, and tpi in G. duodenalis, respectively. Moreover, assemblage A (n = 2) and assemblage E (n = 81) by bg, assemblage A (n = 1) and assemblage E (n = 52) by gdh, and assemblage A (n = 2) and assemblage E (n = 57) by tpi were identified. Multilocus genotyping yielded 29 assemblage E MLGs, which formed 10 subgroups. CONCLUSIONS: To the best of our knowledge, this is the first study to report cattle infected with G. duodenalis in Shanxi, China. Livestock-specific G. duodenalis assemblage E was the dominant assemblage genotype, and zoonotic sub-assemblage AI was also detected in this region.


Assuntos
Giardia lamblia , Giardíase , Humanos , Bovinos , Animais , Giardia lamblia/genética , Tipagem de Sequências Multilocus , Proteínas de Protozoários/genética , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , Genótipo , China/epidemiologia , Prevalência , Fezes/parasitologia , Triose-Fosfato Isomerase/genética , Glutamato Desidrogenase/genética , Filogenia
15.
Parasit Vectors ; 17(1): 155, 2024 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-38528567

RESUMO

BACKGROUND: Cryptosporidiosis and giardiasis are significant parasitic diseases shared between humans and domestic animals. Due to the close contact between humans and domestic animals in rural areas, it is important to consider the potential transmission of zoonotic parasites from infected domestic animals to humans. This investigation aimed to determine the prevalence and molecular characteristics of Cryptosporidium spp. and Giardia duodenalis in domestic animals and villagers. METHODS: A total of 116 fecal samples from villagers and 686 fecal samples from domestic animals in Heilongjiang Province, China, were analyzed for two parasites using nested polymerase chain reaction (PCR) targeting various genetic loci and DNA sequence analysis of the PCR products. RESULTS: By sequence analysis of the SSU rRNA gene, the prevalence of Cryptosporidium in humans was 0.9% (1/116), with one species of C. parvum (n = 1) detected; among domestic animals, the prevalence was 2.6% (18/686), with five species identified: C. suis (n = 7) and C. scrofarum (n = 7) in pigs, C. meleagridis (n = 1) in chickens, C. andersoni (n = 1) in cattle, and C. canis (n = 2) in foxes. C. parvum and C. canis were further subtyped as IIdA19G1 and XXa4 on the basis of gp60 gene. Regarding G. duodenalis, based on the SSU rRNA, bg, gdh, and tpi genes, the prevalence in domestic animals was 5.1% (31/608), with three assemblages identified: A (n = 1) in pigs, D (n = 1) in foxes, and E (n = 27) in geese, cattle, pigs, ducks, and sheep, along with mixed infection of A + E (n = 1) in one pig and B + E (n = 1) in one sheep. No G. duodenalis was detected in humans (0/116). CONCLUSIONS: The present results show that no overlap of subtypes between animals and villagers was found in Cryptosporidium spp. and G. duodenalis, indicating a minor role of domestic animals in infecting humans in this population. However, the presence of zoonotic protozoa in domestic animals highlights the need for special attention to high-risk individuals during close contact with domestic animals.


Assuntos
Criptosporidiose , Cryptosporidium , Giardia lamblia , Giardíase , Humanos , Animais , Bovinos , Ovinos , Suínos , Giardia lamblia/genética , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Cryptosporidium/genética , Animais Domésticos , Raposas , Galinhas , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , China/epidemiologia , Fezes/parasitologia , Prevalência , Genótipo
16.
Parasitology ; 151(4): 351-362, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38305092

RESUMO

Cryptosporidium spp., Giardia intestinalis and microsporidia are unicellular opportunistic pathogens that can cause gastrointestinal infections in both animals and humans. Since companion animals may serve as a source of infection, the aim of the present screening study was to analyse the prevalence of these intestinal protists in fecal samples collected from dogs living in 10 animal shelters in central Europe (101 dogs from Poland and 86 from the Czech Republic), combined with molecular subtyping of the detected organisms in order to assess their genetic diversity. Genus-specific polymerase chain reactions were performed to detect DNA of the tested species and to conduct molecular subtyping in collected samples, followed by statistical evaluation of the data obtained (using χ2 or Fisher's tests). The observed prevalence was 15.5, 10.2, 1 and 1% for G. intestinalis, Enterocytozoon bieneusi, Cryptosporidium spp. and Encephalitozoon cuniculi, respectively. Molecular evaluation has revealed the predominance of dog-specific genotypes (Cryptosporidium canis XXe1 subtype; G. intestinalis assemblages C and D; E. cuniculi genotype II; E. bieneusi genotypes D and PtEbIX), suggesting that shelter dogs do not pose a high risk of human transmission. Interestingly, the percentage distribution of the detected pathogens differed between both countries and individual shelters, suggesting that the risk of infection may be associated with conditions typical of a given location.


Assuntos
Criptosporidiose , Cryptosporidium , Doenças do Cão , Enterocytozoon , Fezes , Giardíase , Microsporidiose , Animais , Cães , Doenças do Cão/parasitologia , Doenças do Cão/epidemiologia , Doenças do Cão/microbiologia , Enterocytozoon/genética , Enterocytozoon/isolamento & purificação , Enterocytozoon/classificação , Cryptosporidium/genética , Cryptosporidium/isolamento & purificação , Cryptosporidium/classificação , Microsporidiose/veterinária , Microsporidiose/epidemiologia , Polônia/epidemiologia , Criptosporidiose/epidemiologia , Criptosporidiose/parasitologia , Fezes/parasitologia , Fezes/microbiologia , República Tcheca/epidemiologia , Giardíase/veterinária , Giardíase/epidemiologia , Giardíase/parasitologia , Prevalência , Giardia/genética , Giardia/isolamento & purificação , Giardia/classificação , Genótipo , Giardia lamblia/genética , Giardia lamblia/isolamento & purificação , Giardia lamblia/classificação , Especificidade de Hospedeiro
17.
Parasitol Int ; 100: 102871, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38403046

RESUMO

Blastocystis sp., Enterocytozoon bieneusi, and Giardia duodenalis are three common zoonotic intestinal parasites, and cattle are important hosts of these three intestinal protozoa. In this study, 1632 fecal samples were collected from dairy farms in Heilongjiang Province, China, and screened for Blastocystis sp., E. bieneusi, and G. duodenalis using polymerase chain reaction. Of these, 149 (9.13%) were positive for three zoonotic pathogens, including 104 (6.40%), 22 (1.35%), and 23 (1.41%) for Blastocystis sp., E. bieneusi, and G. duodenalis, respectively. Based on partial SSU rRNA gene sequencing analysis, 104 positive samples of Blastocystis sp. were found, and a total of nine known subtypes were identified, including ST10 (61), ST3 (18), ST14 (6), ST26 (7), ST24 (3), ST25 (2), ST1 (2), ST5 (2), and ST21 (1). Among these, three subtypes (ST1, ST3, and ST5) were recognized as zoonotic subtypes, and two subtypes (ST10 and ST14) were specific to animals. All 23 Giardia duodenalis-positive samples belonged to assemblage E (n = 23) based on sequenced beta-giardin (bg) and triosephosphate isomerase (tpi) genes. Three known genotypes of E. bieneusi, namely J (n = 9), I (n = 6), and BEB4 (n = 7), were identified by sequence analysis of the internal transcriptional spacer region gene. Our study provides basic data for prevention and control in Heilongjiang Province; however, further research is required to better understand the prevalence and public health significance of these pathogens in the Heilongjiang region.


Assuntos
Cryptosporidium , Enterocytozoon , Giardia lamblia , Giardíase , Microsporidiose , Animais , Bovinos , Giardia lamblia/genética , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , Enterocytozoon/genética , Microsporidiose/epidemiologia , Microsporidiose/veterinária , China/epidemiologia , Genótipo , Fezes/parasitologia , Prevalência , Cryptosporidium/genética
18.
Vet Parasitol ; 327: 110151, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38422710

RESUMO

Rabbits are highly abundant in many countries and can serve as reservoirs of diseases for a diversity of pathogens including the enteric protozoan parasites, Cryptosporidium and Giardia. Both parasites shed environmentally robust environmental stages (oo/cysts) and have been responsible for numerous waterborne outbreaks of diseases. Cryptosporidium hominis and C. parvum are responsible for most infections in humans, while Giardia duodenalis assemblages A and B, cause most human cases of giardiasis. Cryptosporidium cuniculus, the dominant species infecting rabbits, is the only spceies other than C. hominis and C. parvum to have caused a waterborne outbreak of gastritis, which occurred in the United Kingdom in 2008. This review examines the prevalence of Cryptosporidium and Giardia species in rabbits to better understand the public health risks of contamination of water sources with Cryptosporidium and Giardia oo/cysts from rabbits. Despite the abundance of C. cuniculus in rabbits, reports in humans are relatively rare, with the exception of the United Kingdom and New Zealand, and reports of C. cuniculus in humans from the United Kingdom have declined substantially since the 2008 outbreak. Subtyping of C. cuniculus has supported the potential for zoonotic transmission. Relatively few studies have been conducted on Giardia, but assemblage B dominates. However, improved typing methods are required to better understand the transmission dynamics of Giardia assemblages in rabbits. Similarly, it is not well understood if pet rabbits or contaminated water are the main source of C. cuniculus infections in humans. Well-planned studies using high-resolution typing tools are required to understand the transmission dynamics better and quantify the public health risk of Cryptosporidium and Giardia from rabbits.


Assuntos
Criptosporidiose , Cryptosporidium , Cuniculidae , Cistos , Giardia lamblia , Giardíase , Doenças dos Roedores , Coelhos , Humanos , Animais , Giardíase/epidemiologia , Giardíase/veterinária , Giardíase/parasitologia , Giardia , Criptosporidiose/parasitologia , Zoonoses/parasitologia , Água/parasitologia , Fezes/parasitologia , Cistos/veterinária
19.
Vet Parasitol ; 327: 110139, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38295676

RESUMO

Giardia duodenalis is one of the most prevalent enteric parasites of dogs. Point-of-care antigen tests (POC) are rapid and do not require additional equipment, or a specialised diagnostic laboratory. The aim of this study was to compare diagnostic tests available in veterinary practices and in a diagnostic laboratory for the detection of G. duodenalis on a cohort of group-housed dogs from New South Wales, Australia. Two different POC tests were used for the detection of G. duodenalis. Laboratory tests used were the multiplexed-tandem PCR panel (MT-PCR) that includes detection of G. duodenalis DNA, and two reference tests (an in-house TaqMan real-time PCR and a direct immunofluorescence assay, DFA). Canine faecal samples (n = 40) were tested simultaneously for the detection of G. duodenalis. Using either DFA or TaqMan real-time PCR as reference tests, 77.5% (31/40) and 82.5% (33/40) of dogs tested positive, respectively. Agreement (Kappa) between the DFA and TaqMan real-time PCR was 0.84 (95% CI 0.64 to 1.00). There was substantial G. duodenalis test outcome agreement between the two POC tests, Kappa = 0.75. Combining the two POC tests yielded 77% sensitivity and 100% specificity with DFA as reference, and for TaqMan real-time PCR it was 73% sensitivity and 100% specificity. The MT-PCR was in excellent agreement with each reference test, DFA or TaqMan real-time PCR. Due to the high specificity of both POC tests, they can be confidently used as rule-in diagnostics. Confirmatory testing that detects different biological parameters such as DNA, e.g. PCR (inc. MT-PCR), should be implemented before concluding that a dog is negative for the presence of G. duodenalis.


Assuntos
Doenças do Cão , Giardia lamblia , Giardíase , Humanos , Cães , Animais , Giardia lamblia/genética , Sistemas Automatizados de Assistência Junto ao Leito , Reação em Cadeia da Polimerase Multiplex/veterinária , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Antígenos de Protozoários/genética , DNA , Giardíase/diagnóstico , Giardíase/veterinária , Giardíase/parasitologia , Fezes/parasitologia , Doenças do Cão/diagnóstico , Doenças do Cão/parasitologia
20.
J Struct Biol ; 216(1): 108064, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38280689

RESUMO

The inner structure of the flagella of Giardia intestinalis is similar to that of other organisms, consisting of nine pairs of outer microtubules and a central pair containing radial spokes. Although the 9+2 axonemal structure is conserved, it is not clear whether subregions, including the transition zone, are present in the flagella of this parasite. Giardia axonemes originate from basal bodies and have a lengthy cytosolic portion before becoming active flagella. The region of the emergence of the flagellum is not accompanied by any membrane specialization, as seen in other protozoa. Although Giardia is an intriguing model of study, few works focused on the ultrastructural analysis of the flagella of this parasite. Here, we analyzed the externalization region of the G. intestinalis flagella using ultra-high resolution scanning microscopy (with electrons and ions), atomic force microscopy in liquid medium, freeze fracture, and electron tomography. Our data show that this region possesses a distinctive morphological feature - it extends outward and takes on a ring-like shape. When the plasma membrane is removed, a structure surrounding the axoneme becomes visible in this region. This new extra-axonemal structure is observed in all pairs of flagella of trophozoites and remains attached to the axoneme even when the interconnections between the axonemal microtubules are disrupted. High-resolution scanning electron microscopy provided insights into the arrangement of this structure, contributing to the characterization of the externalization region of the flagella of this parasite.


Assuntos
Axonema , Giardia lamblia , Giardia lamblia/ultraestrutura , Microtúbulos/metabolismo , Flagelos/metabolismo , Microscopia Eletrônica de Varredura
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